Methods and compositions for modulating immune response and for the treatment of inflammatory disease

ABSTRACT

Compositions and methods for treating inflammatory disease and for modulating the immune system are described. The compositions and methods pertain to the use of a combination of N,N-dimethylglycine and a component derived from the mussel  Perna canaliculus  to achieve synergistic anti-inflammatory and immunomodulating effects.

BACKGROUND OF INVENTION

[0001] This invention relates to the use of N,N-dimethylglycine (DMG) incombination with at least one component derived from Perna canaliculusto modulate immune responses and to treat inflammatory diseases in manor animals.

[0002] N′ N-dimethylglycine (DMG), which is a tertiary amino acid, is anintermediary metabolite found in low levels in many foods. It isproduced in the body from choline and has been used as a non-fuelnutrient. The physiological and potential therapeutic effects ofN,N-dimethylglycine have been evaluated in recent years. References inthe literature pertaining to N,N-dimethylglycine and its potential usesinclude the following: Graber et al., 1981, J. of Infectious Diseases143:101-105 (DMG stimulation of both humoral and cellular immunity);Kendall and Graber, U.S. Pat. No. 4,385,068 (discusses DMG alleviationof the effects of excess radiation on the immune system); the 1980Pacific Slope Biochemical Conference, a paper entitled “Decrease ofLactic Acid Concentration in Blood of Animals GivenN,N-Dimethylglycine”; March, 1982 issue of Equine Practice, an articleentitled “Effect of a Nutritional Supplement ContainingN,N-Dimethylglycine (DMG) on the Racing Standardbred” (DMG increasedoxygen utilization and thereby decreased lactic acid levels in animalsunder extreme stress); November-December, 1982 issue of Canine Practice,an article entitled “A Clinical Evaluation of N,N-Dimethylglycine (DMG)and Diisopropylammonium Dichloroacetate (DIPA) on the Performance ofRacing Greyhounds”; February, 1987 issue of Let's Live magazine, anarticle entitled “Dimethylglycine Update, New Studies Confirm DMGImproves Health” (DMG as an immunomodulating agent); February, 1987issue of Health Consciousness, an article entitled “N,N-Dimethylglycineand the Immune Response” (DMG as an immunomodulating agent); 1987 ASMAnnual Meeting, a paper entitled “The Effect of DMG on the ImmuneResponse of Rabbits” (DMG as an immunomodulating agent); Jun. 27, 1987issue of The Blood Horse, an article entitled “DMG, Properties andProprieties” (in humans, DMG stimulated B-cells to produce much higherantibody responses, and DMG enhanced the activity of T cells andmacrophages); Kendall et al., U.S. Pat. No. 4,994,492 (treatment ofmelanoma using DMG); Kendall and Lawson, U.S. Pat. No. 5,026,728(treatment of arthritis and inflammation using DMG); Kendall and Lawson,U.S. Pat. No. 5,118,618 (use of DMG to enhance antibody production).

[0003] The use of preparations made from Perna canaliculus fortherapeutic effect extends back at least twenty-five years since themid-seventies. The nutritional and therapeutic properties offreeze-dried Perna canaliculus in alleviating the symptoms of arthritiswere reported in Croft, 1979, Relief From Arthritis (Thorsons PublishingGroup, Rochester, Vt.). Studies in both animal and human experimentshave given mixed results, but indicate overall that certain componentsin the mussels were potentially helpful in relieving inflammation andpain associated with arthritis. Certain of these studies are as follows:McFarlane, June 1975, New Zealand Medical Journal, pg. 569 (the resultsof a study of the effects of a Perna canaliculus product called“Seatone” on a small group of arthritic individuals were reported);Rainsford and Whitehouse, 1980, Arzeim-Forsch, pg. 2128-2131 (afreeze-dried powdered preparation of whole mussel which ws given orallyto rats showed some modest anti-inflammatory activity in an inducedrheumatoid arthritis model); Miller and Ormrod, 1980, The New ZealandMedical Journal 92:187 (a crude fraction of Perna canaliculus had amarked anti-inflammatory effect when administered by intra peritonealinjection in rats); Couch et al., 1982, The New Zealand Medical Journal95:803 (a protein containing fraction of Perna canaliculus had ananti-inflammatory effect); Caughey et al., 1983, European Journal ofRheumatology and Inflammation 6:197 (in a human study involvingforty-seven rheumatoid arthritic patients, no improvement was observedin patients given mussel extract as compared to patients taking theanti-inflammatory drug naproxen); Gibson et al., 1980, Practitioner224:955 (93:111 (in human trials for both osteoarthritis and rheumatoidarthritis, significant benefits were achieved after three months on aPerna canaliculus preparation, including reduced pain and stiffness andimprovement on functional tests); Audeval and Bouchart, 1986, GazetteMedicale 93:111 (in human trials for osteoarthritis, forty percent ofsubjects showed significant improvement after treatment with a Pernacanaliculus preparation); Miller et al., 1993, Agents Actions 38:139 (anaqueous fraction of Perna canaliculus inhibited experimentally inducedinflammation in rats); Whitehouse et al., 1997, Inflammopharmacology (alipid extract of Perna canaliculus exhibited a dose relatedanti-inflammatory activity).

SUMMARY OF THE INVENTION

[0004] In accordance with this invention, it has been discovered that acombination of N,N-dimethylglycine (DMG) and a component derived fromPerna canaliculus (referred to herein as Perna canaliculus extract(PCE)) has a synergistic effect in the treatment of inflammatorydisease. In particular, it has been discovered that the combination hasa synergistic effect in the treatment of autoimmune disease, includinglupus erythmatosus, and in the modulation of immune responses toinflammatory disease. Also in accordance with this invention it has beendiscovered that the aberrant immune response profiles in humans withrheumatoid arthritis or osteoarthritis may be beneficially altered bythe administration of a combination of DMG and PCE.

[0005] Treatment, as used herein, pertains to the therapeuticadministration of the compounds of the invention for the prevention,amelioration, or cure of disease.

[0006] Inflammatory diseases that may be treated in accordance with thisinvention include systemic autoimmune diseases such as, for example,lupus erythmatosus, rheumatoid arthritis, and multiple sclerosis, andorgan specific autoimmune diseases such as, for example, myastheniagravis, Grave's disease, Hashimoto's thyroiditis, Crohn's disease,autoimmune hemolytic anemias, insulin dependent diabetes mellitus,glomerulonephritis, and rheumatic fever. Other inflammatory diseasesthat may be treated in accordance with this invention include otherinflammatory arthritic conditions such as osteoarthritis and goutyarthritis, as well as other inflammatory conditions such asconjunctivitis, dermatitis, bronchitis, rhinitis etc., brought about byinjury, allergies, infections, microorganisms, trauma, or physical orchemical agents. Additionally, the treatment of inflammatory aspects ofasthma is also contemplated as part of this invention.

[0007] One aspect of the invention is a composition comprising acombination of a DMG component and at least one PCE component. The PCEcomponent may be any therapeutically active component derived from theflesh of the mussel or its organs, which are suitable for use in thepreparation of food supplements or pharmaceutical preparations. Theinvention contemplates the use of unrefined components of the mussel,such as whole mussel, or any therapeutically active extracts thereof. Asused herein, a therapeutically active PCE component according to thisinvention is a component that is effective in reducing inflammation inanimals or humans or that has immunomodulating effects in humans oranimals. For example, several animal models for arthritis that are wellknown in the art may be used to determine therapeutic activity of a PCEcomponent. Such models include, for example, collagen induced arthritisin rats or mice, carrageenan-induced inflammatory oedema in rats asdescribed in Miller et al., 1980, New Zealand Medical Journal 92:667,and arthritis induced in rats and mice by the injection of adjuvantsprepared from dried Mycobacterium tuberculosis as described inWhitehouse, et al., 1997, Inflammopharmacology 5:237-246. Therapeuticactivity of a PCE component may also be determined by evaluating whetherthe PCE component is effective for reducing anti-nuclear antibodies,particularly anti-ssDNA antibodies or anti-dsDNA antibodies, in humanswith SLE or in the mouse model for SLE, as described in the examples. Inparticular, the PCE may be a preparation of whole mussel which has beenfreeze-dried and ground such as, for example, the Sea Mussel productavailable from FoodScience Corporation, Essex Junction, Vt. or the Pernaproduct available from DaVinci Laboratories, Essex Junction, Vt.Additionally, therapeutically active extracts of Perna canaliculus mayalso be employed. Such therapeutically active extracts have beendescribed as follows: Macrides and Kalafatis, PCT/AU95/00485, WO96/05164 (a purified lipid extract); Whitehouse et al., 1997,Inflammopharmacology 5:237-246 (a purified lipid extract); Kosuge andSugiyama, U.S. Pat. No. 4,801,453 (stabilized mussel extract); Couch etal., 1982, The New Zealand Medical Journal 95:803 (a protein fraction);Miller et al., 1993, Agents Actions 38:139 (an aqueous fraction).

[0008] The DMG component may be N,N-dimethylglycine or apharmacologically acceptable salt thereof. A composition containing bothDMG and PCE may be formulated as a pharmaceutical or dietary supplementusing techniques that are well known in the art. A preferred formulationof the composition is a formulation for oral administration.

[0009] Another aspect of this invention is a method of treatinginflammatory disease comprising administering to an animal a combinationof both a DMG component and at least one PCE component. As used herein,the term animal includes, but is not limited to, mammals. Preferredmammals include humans, horses, farm animals and household pets. The DMGand PCE components may be administered via the same route or they may beadministered via different routes. For example, the DMG and PCE may bothbe administered orally, either simultaneously or at different times. Apreferred route for administration of PCE, particularly ground wholemussel, is via oral administration. A preferred route for theadministration of DMG is via oral administration, particularly as anadmixture with water. DMG or PCE can also be administered byintramuscular injection, intra peritoneal injection, parenteraladministration, etc.

[0010] The DMG and PCE used in this invention can be employed inadmixture with conventional excipients, i.e. pharmaceutically acceptableorganic or inorganic carrier substances suitable for parenteral, enteral(e.g., oral) application which do not deleteriously affect the activecompound. Suitable pharmaceutically acceptable carriers include but arenot limited to water, salt solutions, alcohols, gum arabic, vegetableoils, benzyl alcohols, polyethylene glycols, gelatine, carbohydratessuch as lactose, amylose or starch, magnesium sterate, talc, silicicacid, viscous paraffin, perfume oil, fatty acid monoglycerides anddiglycerides, pentaerythritol fatty acid esters, hydroxymethylcellulose, polyvinyl pyrrolidone, etc. The pharmaceuticalpreparations can be sterilized and, if desired, mixed with auxiliaryagents such as, for example, lubricants, preservatives, stabilizers,wetting agents, emulsifiers, salts for influencing osmotic pressure,buffers, coloring agents, flavoring agents and/or aromatic substancesand the like which do not deleteriously affect the active compound.Other pharmaceutically acceptable carriers include aqueous solutions,non toxic excipients, including salts, preservatives, buffers and thelike, as described for instance, in Remington 's PharmaceuticalSciences, 18th ed. (1990), Mack Publishing Co., Easton, Pa., thecontents of which are hereby incorporated by reference. The pH and exactconcentration of the various components of the pharmaceuticalcomposition are adjusted according to routine skills in the art. SeeGilman et al. (eds.) (1990) Goodman and Gilman 's: The PharmacologicalBases of Therapeutics, 8th ed., Pergamon Press. The DMG and/or PCE canalso be combined where desired with other active agents, including PCEor DMG respectively.

[0011] The pharmaceutical compositions are preferably prepared andadministered in dose units. Solid dose units are tablets, capsules andsuppositories. For treatment of a subject, depending on activity of thecompound, manner of administration, nature and severity of the disorder,age and body weight of the patient, different daily doses are necessary.Under certain circumstances, however, higher or lower daily doses may beappropriate. The average daily dosage of the compounds of thisinvention, when used to treat inflammatory disease or to modulate theimmune response are typically the following: for DMG, the average dailydose is generally about 1 to about 500 mg/kg/day, preferably about 10 toabout 100 mg/kg/day, more preferably about 2 to about 30 mg/kg/day; forPCE, when the PCE comprises ground whole mussel, the average daily doseis generally about 1 to about 500 mg/kg/day, preferably about 10 toabout 100 mg/kg/day, more preferably about 20 to about 60 mg/kg/day. ForPCE components other than ground whole mussel, the average daily dosemay vary but can be readily determined by one of skill in the art. Thedaily dosage may be administered as a single dose per day or as aplurality of divided doses per day. The daily dosage may also beadministered on a non-daily basis, such as for example, every second orthird day, provided an average daily dose as described above isachieved.

[0012] The combination PCE and DMG can be administered concurrently oralternately with other therapeutic treatments conventionally employed totreat inflammatory disease.

[0013] For example, for the treatment of lupus erythmatosus, rheumatoidarthritis, and osteoarthritis, PCE and DMG may be administered inconjunction anti-inflammatory agents, including both coticosteroidagents such as, for example, prednisone or methylprednisolone andnon-steroidal anti-inflammatory agents such as, for example, salicylatesand ibuprofen. For the treatment of lupus erythmatosus, for example, thecompounds of this invention may also be administered in conjunction withanti-malarial drugs including, for example, hydroxychloroquinone or inconjunction with cytotoxic chemotherapies including, for example,azathioprine and cyclophosphamide.

[0014] Another aspect of this invention is a method of treating systemiclupus erythmatosus comprising administering a combination of DMG andPCE. In a preferred embodiment, the administration of the combinationreduces the serum concentration of anti-nuclear antibodies such as, forexample, anti-nuclear antibodies that bind to chromatin, nucleosomes,histone/DNA complexes, histones, double-stranded DNA (dsDNA),single-stranded DNA (ssDNA), double-stranded RNA (dsRNA),single-stranded RNA (ssRNA), and transfer-RNA (tRNA). Preferably, theserum concentration of anti-ssDNA and/or anti-dsDNA antibodies arereduced.

[0015] Another aspect of this invention is a method modulating theimmune response in an animal, particularly an animal with an autoimmunedisease or other inflammatory disease by the administration of acombination of DMG and PCE, wherein any of the following effects on theimmune response, or any combination thereof, are achieved: a decrease inthe serum levels of interleukin 6 (il-6), a decrease in the serum levelsof interleukin 10 (il-10), an increase in the serum levels of tumornecrosis factor-α (TNF-α), or a decrease in the concentration of CD8⁺lymphocytes in the blood. These effects typically counteract aberrantimmune profiles observed in inflammatory disease. A preferred effect isa decrease in the serum levels of il-10 or a decrease in CD8⁺lymphocytes in the blood.

[0016] Another aspect of this invention is a kit containing acombination of DMG and PCE formulations. Particularly preferred is kitcontaining both DMG and PCE formulations that are useful for oraladministration, preferably as dietary supplements. A kit would typicallyprovide for an average daily dose of PCE or DMG as described above.

BRIEF DESCRIPTION OF THE FIGURES

[0017]FIG. 1 is a graph showing the weight gain comparisons for controlmice, and for mice treated with DMG, PCE, or a combination of DMG andPCE.

[0018]FIG. 2 is a graph comparing cytokine concentrations, includingil-4, il-6, il-10, and TNF-α, in control mice and in mice treated withDMG, PCE, or a combination of DMG and PCE.

[0019]FIG. 3 is a graph comparing lymphocyte population levels,including lymphocytes having CD4, CD8, and CD19 markers, in control miceand in mice treated with DMG, PCE, or a combination of DMG and PCE.

[0020]FIG. 4 is a graph comparing concentrations of anti-double strandedDNA antibodies in control mice and in mice treated with DMG, PCE, or acombination of DMG and PCE. Two experiments, (#1) and (#2), wereconducted.

[0021]FIG. 5 is a graph comparing concentrations of an anti-singlestranded DNA antibodies in control mice and in mice treated with DMG,PCE, or a combination of DMG and PCE.

[0022]FIG. 6 is a graph comparing lymphocyte populations in healthyhuman subjects with lymphocyte populations in human subjects withosteoarthritis or rheumatoid arthritis. Cell populations that weremeasured include total T cells (T Cells), helper T cells (Th) (CD4⁺),cytotoxic T cells (Tc) (CD8⁺), B cells (B) (CD19⁺), and natural killercells (NK). Within a subpopulation, columns with different superscripts(i.e. a and b) are significantly different (p≦0.01).

[0023]FIG. 7 is a graph comparing cytokine concentrations in healthyhuman subjects with cytokine concentrations in human subjects withosteoarthritis or rheumatoid arthritis. Within a subpopulation, columnswith different superscripts (i.e. a and b) are significantly different(p<0.01).

[0024] Without further elaboration, it is believed that one skilled inthe art can, using the preceding description, utilize the presentinvention to its fullest extent. The following preferred specificembodiments are, therefore, to be construed as merely illustrative, andnot limitative of the remainder of the disclosure.

EXAMPLES

[0025] In the following examples, all parts and percentages are byweight unless otherwise indicated.

Example 1 Preparation of a Therapeutically Active PCE Component

[0026] A therapeutically active formulation of Perna canaliculus isprepared by freeze-drying the flesh of the mussel and grinding it into apowder. The product is formulated into capsules with the excipients ofalfalfa, cellulose and magnesium stearate.

Example 2 Treatment of Systemic Lupus Erythmatosus with a Combination ofDMG and PCE Extract

[0027] Systemic lupus erythmatosus (SLE) is considered to be theprototypic autoimmune disease. SLE is characterized by various degreesof severity with mild forms resulting in minor skin rash and joint painand severe forms resulting in glomerulonephritis, immune complexdisease, severe lymph node enlargement, and multiple organ failure. SLEis further characterized by the development of auto-antibodies againstcytoplasmic and nuclear components. In addition to clinical symptoms,the presence of anti-nuclear antibodies is a hallmark of the disease,and its presence is an important indication in the diagnosis of thedisease (Tan, 1989, Adv. Immunology 44:93). MRI-lpr mice are an acceptedand well documented model for SLE (Cohen and Eisenberg, 1991, Ann. Rev.Immunol. 9:203; Kotzin, 1996, Cell 85:303). These mice display ahomozygous genetic defect in the Fas-TNF receptor related to apoptosis.Fas protein is an important pathway for the induction of apoptosis andthe Fas protein is responsible for lymphocyte removal. These miceproduce a broad spectrum of auto-antibodies against nuclear andcytoplasmic proteins, and the MRI-lpr mice die from classical lupus-likesymptoms.

[0028] MRI-lpr mice were used to study the effects of a combination ofDMG and PCE on components of the immune system and on the progression ofthe SLE disease. Forty MRI-lpr mice were divided into four groups of tenmice each: a control group, a PCE treated group, a DMG treated group,and a PCE-DMG treated group. The control group was given standard mousechow and water. The DMG group was given standard mouse chow and watermixed with 34 mg/L of DMG (AANGAMIK® DMG, FoodScience Corporation, EssexJunction, Vt.). The PCE group was given a PCE-mouse chow mix and water.The PCE-DMG combination group was given a PCE-mouse chow mix and DMGwater. The PCE supplement used in the studies was a product containingfreeze-dried, ground whole Perna canaliculus mussel (Perna, FoodScienceCorporation, Essex Junction, Vt.). The mice were given food and water adlibitum. It is estimated that the mice, with a size range of 30-50grams, consumed about 1-3 mg/day of DMG and 2-6 mg/day of PCE. The micewere weighed weekly and 100 ul of blood was collected via orbital bleedon a weekly basis. The collected blood was centrifuged and the serum wasrecovered and stored at −20° C. for analysis at the end of the twelveweek experiment. After twelve weeks, the mice were sacrificed and thespleens, kidneys, and livers were harvested. When the mice weresacrificed, blood was also collected via a cardiac stick. The serum wascollected and stored as described above. The spleens were processed andthe cells recovered were quantified by flow cytometry using fluorescentanti-CD4, anti-CD8 and anti-CD19 antibodies (Pharmingen, San Diego,Calif.). Cytokine levels in the serum and anti-single stranded anddouble stranded DNA antibody levels in the serum were determined usingan enzyme linked immunosorbent assay (ELISA).

[0029] Animals whose diets were supplemented with PCE alone or theDMG-PCE combination experienced a higher weight gain than control or DMGtreated animals, indicating that the mice treated with the combinationwere not adversely affected by the treatment (FIG. 1). A significantdecrease in CD8⁺ T cells was noted with animals fed the standard DMG-PCEcombination, with a trend toward lower levels in animals fed only PCE(FIG. 3). A trend toward lower CD19⁺ B cell percentages was alsoobserved with DMG-PCE treated animals, while animals treated with PCEalone had CD19⁺ B cell levels comparable to those of controls. No effectof DMG alone on CD8⁺T-cell or CD19⁺ B-cell levels could be detected(FIG. 3).

[0030] The effects on cytokine production in mice treated with theDMG-PCE combination was also examined. As shown in FIG. 2, animalstreated with the DMG-PCE combination showed significantly lower levelsof serum il-6 than either control animals or animals treated with DMGalone. Although animals treated with either DMG or PCE alone hadsignificantly lower levels of il-6 than control animals, animals treatedwith a combination of the two had even lower levels. DMG-PCE treatedanimals also showed significantly lower levels of serum il-10 thaneither control animals or animals treated with either DMG or PCE alone.These changes in cytokine serum levels are indicative of a shift from aTh2 response to a Th1 response. The significant rise in serum levels oftumor necrosis factor (TNF) in DMG-PCE treated animals is alsoindicative of this shift from a Th2 response to a Th1 response.

[0031] Treatment of animals with DMG alone resulted in higher levels ofIL-10 than in controls, indicating a shift toward a Th2 response.Although DMG is a known enhancer of antibody production, it is not knownwhether the enhanced antibody production observed representspathological IgG2a production. Levels of serum il-10 and TNF-α werecomparable in control animals and in animals treated with PCE alone.Therefore, DMG-PCE treatment appeared to shift the lymphocyte responsefrom a Th2 response to a Th1 response, with a concomitant decrease inexcess antibody production, including anti-nuclear antibody productionas discussed below.

[0032] Levels of anti-nuclear antibodies were also examined in controland treated mice. After eleven weeks of treatment, anti-dsDNA antibodylevels in the serum of DMG-PCE treated animals were significantly lowerthan in control animals or in animals treated with DMG alone or PCEalone (FIG. 4). Even more significantly, anti-ssDNA antibody levels inthe serum of DMG-PCE treated animals were in tandem with levels incontrol animals until the 7th week, when these serum concentrations werefound to be lower (FIG. 5). This significant difference was observed tocontinue until animals were euthanized after the 11th week. Serum levelsof anti-ssDNA antibodies in control animals and in animals treated withPCE alone continued to increase throughout the course of treatment. Bythe 11th week, serum levels in animals treated with DMG alone appearedto be leveling out.

Example 3 Comparison of Lymphocyte Populations in Healthy Human Subjectswith Lymphocyte Populations in Human Subjects with Osteoarthritis orRheumatoid Arthritis.

[0033] Lymphocyte populations were measured and compared in healthyhuman subjects versus human subjects with osteoarthritis or rheumatoidarthritis (FIG. 6). Lymphocyte populations measured included total Tcells (T Cells), helper T cells (Th) (CD4⁺ cells), cytotoxic T cells(Tc) (CD8⁺ cells), B cells (B), and natural killer cells (NK). The cellpopulations were identified by external staining of whole bloodlymphocytes using the following antibodies: anti-CD3 antibodies fortotal T cells; anti-CD4 antibodies for helper T-cells; anti-CD8antibodies for cytotoxic T; anti-CD5 and anti-CD19 antibodies for Bcells; anti-CD56 antibodies for natural killer cells; and anti-CD45antibodies for monocytes. The antibodies were obtained from eitherCaltag Corp. or Coulter Corp. and were tagged for analysis on an EPICs751 flow cytometer (Coulter F1 ) with a Cicero acquisition module andcyclops analysis software (Cytomation, Calif.). As shown by the data inFIG. 6, a CD4⁺/CD8⁺ T cell switch was observed in both diseases whencompared to healthy controls, with a significant decrease in CD4⁺ cellsin subjects with either disease and a significant increase in CD8⁺ cellsin subjects with either disease.

Example 4 Comparison of Cytokine Concentrations in Healthy HumanSubjects with Cytokine Concentrations in Human Subjects withOsteoarthritis or Rheumatoid Arthritis.

[0034] Cytokine levels were measured and compared in healthy humansubjects versus human subjects with osteoarthritis or rheumatoidarthritis (FIG. 7). Cytokines measured included gamma interferon,interleukin 1—beta, interleukin 2, interleukin 4, interleukin 6,interleukin 10 and TNF-α. Cytokines were measured using specific ELISAkits according to the manufacturers instructions (Immunotech, France).Interleukin 6 levels were significantly higher in subjects with eitherdisease than for control subjects. TNF-α levels were significantlyincreased in subjects with rheumatoid arthritis compared to normalcontrols, but were not increased in patients with osteoarthritis.

[0035] The preceding examples can be repeated with similar success bysubstituting the generically or specifically described reactants and/oroperating conditions of this invention for those used in the precedingexamples.

[0036] Without further elaboration, it is believed that one skilled inthe art can, using the preceding description, utilize the presentinvention to its fullest extent. The preceding preferred specificembodiments are, therefore, to be construed as merely illustrative, andnot limitative of the remainder of the disclosure in any way whatsoever.

[0037] The entire disclosure of all patent applications, patents, andpublications cited herein are hereby incorporated by reference.

[0038] From the foregoing description, one skilled in the art can easilyascertain the essential characteristics of this invention and, withoutdeparting from the spirit and scope thereof, can make various changesand modifications of the invention to adapt it to various usages andconditions.

What is claimed is:
 1. A method of treating a subject for inflammatorydisease comprising administering to the subject a DMG component and atleast one PCE component.
 2. A method according to claim 1, wherein thesubject has an autoimmune disease.
 3. A method according to claim 2,wherein the autoimmune disease is lupus erythmatosus.
 4. A methodaccording to claim 3, wherein a reduction in serum concentration of atleast one anti-nuclear antibody is achieved.
 5. A method according toclaim 4, wherein the anti-nuclear antibody is an anti-dsDNA antibody oran anti-ssDNA antibody.
 6. A method according to claim 3, wherein atleast one PCE component is freeze-dried ground whole mussel.
 7. A methodof modulating the immune response profile in a subject comprisingadministering to the subject a DMG component and at least one PCEcomponent.
 8. A method according to claim 7, wherein the modulationresults in a decrease in serum il-6 concentrations, a decrease in serumil-10 concentrations, an increase in serum TNF-α concentrations, or adecrease in CD8⁺ cells in the blood.
 9. A method according to claim 7,wherein the modulation results in a decrease in serum il-10concentration and a decrease in CD8⁺ cells in the blood.
 10. A methodaccording to claim 7, wherein the modulation results in a decrease inCD8⁺ cells.
 11. A method according to claim 10, wherein the subject hasan autoimmune disease.
 12. A composition comprising a DMG component andat least one PCE component.
 13. A composition according to claim 12,wherein at least one PCE component comprises freeze-dried ground wholemussel.
 14. A composition according to claim 12 which is suitable foruse as a dietary supplement.
 15. A kit comprising a DMG formulation anda PCE formulation, wherein the PCE formulation comprises at least onePCE component.
 16. A kit according to claim 15, wherein the DMGformulation and the PCE formulation are suitable for use as dietarysupplements.
 17. A kit according to claim 15, wherein at least one PCEcomponent comprises ground freeze-dried whole mussel.
 18. A kitaccording to claim 15, wherein the DMG formulation is suitable foradministration as an admixture with water.